Enhanced Ca2+ response and stimulation of prostaglandin release by the bradykinin B2 receptor in human retinal pigment epithelial cells primed with proinflammatory cytokines

Abstract Kallikrein, kininogen and kinin receptors are present in human ocular tissues including the retinal pigment epithelium (RPE), suggesting a possible role of bradykinin (BK) in physiological and/or pathological conditions. To test this hypothesis, kinin receptors expression and function was investigated for the first time in human fetal RPE cells, a model close to native RPE, in both control conditions and after treatment with proinflammatory cytokines. Results showed that BK evoked intracellular Ca 2+ transients in human RPE cells by activating the kinin B 2 receptor. Pretreatment of the cells with TNF-α and/or IL-1β enhanced Ca 2+ response in a time- and concentration-dependent additive manner, whereas the potency of BK and that of the selective B 2 receptor antagonist, fasitibant chloride, both in the nanomolar range, remained unaffected. Cytokines have no significant effect on cell number and viability and on the activity of other GPCRs such as the kinin B 1 , acetylcholine, ATP and thrombin receptors. Immunoblot analysis and immunofluorescence studies revealed that cytokines treatment was associated with an increase in both kinin B 2 receptor and COX-2 expression and with the secretion of prostaglandin E 1 and E 2 into the extracellular medium. BK, through activation of the kinin B 2 receptor, potentiated the COX-2 mediated prostaglandin release in cytokines-primed RPE cells while new protein synthesis and prostaglandin production contribute to the potentiating effect of cytokines on BK-induced Ca 2+ response. In conclusion, overall data revealed a cross-talk between the kinin B 2 receptor and cytokines in human RPE in promoting inflammation, a key feature in retinal pathologies including diabetic retinopathy and macular edema.